Many kinds of mRNA are not translated immediately after transcription, but rather transported to specific subcellular compartments and then start protein synthesis. Regulation of D-actin mRNA trafficking and localization are mediated by a specific sequence ('zipcode') in the RNA, as well as several RNA binding proteins such as zipcode binding protein, ZBP1. Localization of D-actin mRNA is postulated to have a fundamental role in cell motility, tissue differentiation, and development of organs. With recent technical breakthroughs in fluorescent labeling in vivo, the dynamic motions of mRNA and RNA-protein interactions can now be investigated in high spatial and temporal resolutions. Furthermore, with novel imaging methods that improve the sensitivity, in situ visualization of mRNA localization processes at a single molecular level has become routine performance. This program aims to study the localization of endogenous D-actin mRNA in cells and tissues from a transgenic animal model. Recently MS2 mouse model that includes multiple MS2 stem-loops in the 3'UTR region of D-actin mRNA was constructed in Singer laboratory. By associating the MS2 stem-loops with MS2-GFP fusion proteins, it is possible to track the molecule with high signal-to-noise ratio. Specifically single-particle tracking based on the numerical finding of the centroid and fluorescent recovery after photobleaching (FRAP) will enable us to study the nature of mRNA transport in detail. Ultimately the cycles of mRNA movements will be examined in nanometer resolutions to correlate with the translation activities. In order to investigate the physiological significance of D-actin mRNA localization in a tissue microenvironment, tissue sections and the whole organs of living MS2 mouse will be imaged. Multiphoton microscopy will be employed, because of the well-known advantages in tissue studies: reduced out-of-focus photodamage, 3-D optical sectioning, and use of near-infrared laser beam that permits deeper penetration into tissue. Studying D-actin mRNA localization in the context of multicellular interactions in living tissue will shed an insight on human disease processes such as cancer metastasis, and pathological neuronal development.